Development of a Bispecific Antibody Based Immunotherapy for Triple Negative Breast Cancer. SUMMARY/ABSTRACT A promising and versatile approach to cancer treatment is the use of bispecific antibodies (bsAb) for immune cell redirection. In the two clinically approved therapies that pioneered this strategy, a bsAb drug acts as a molecular bridge between a T cell (via CD3 binding) and a tumor cell (via binding of a tumor associated antigen, or TAA). Simultaneous binding of both targets results in T cell mediated cytolysis of the tumor cell. The impressive success of this strategy has led to the development of similar bsAb immunotherapies for a wide variety of both hematological and solid cancers, resulting in over 15 drugs in active clinical trials. Triple negative breast cancer (TNBC) is an invasive breast cancer defined by a lack of the three distinct TAA required for available targeted breast cancer therapies. The lack of a robust TAA for TNBC has resulted in a dearth of modern targeted therapies and poor patient outcomes, and therefore development of novel TAA-targeted drugs for TNBC is a significant unmet medical need. OncoTAb Inc has developed a proprietary antibody (TAB 004) that recognizes a tumor specific variant of MUC1 (tMUC1). This antibody serves as the basis of OncoTAb?s Agkura Personal Score diagnostic test and has high specificity for a broad number of TNBC cell lines. In this proposal, we will develop and test a bsAb based immunotherapy for TNBC derived from the TAB 004 antibody. Historically, development of novel bsAb immunotherapies relies on the use of antibody fragments or complicated post-processing and purification, both of which can reduce stability, hinder manufacturability and prolong development. In order to alleviate these development hurdles, Dualogics, LLC has developed a proprietary bsAb platform, called OrthoMab, that retains the stability and manufacturability of native antibodies and is compatible with existing antibody sequences. Using the OrthoMab platform, we can rapidly generate a suite of bsAb molecules with varied size, binding valency, and geometric orientation; all of which have been shown to modulate the efficacy of bsAb immunotherapies. In Specific Aim 1 of this proposal we will generate and biophysically characterize three bsAb specific for tMUC1 and CD3, each with unique pharmacokinetic and functional properties (Ryan Hallett, Dualogics LLC). In Aim 2, we will measure specificity of each bsAb for TNBC cell lines and demonstrate targeted cytolysis of tumor cells by human PBMCs (Lloye Dillon, OncoTAb Inc.). In Aim 3, we will assess the pharmacokinetic properties of our top performing bsAb to determine dosing and evaluate efficacy in a TNBC xenograft mouse model (Ru Zhou, UNC-Charlotte). These experiments will validate T cell redirection by tMUC1 as a viable strategy for TNBC treatment and establish the basis of a phase II study to pursue further clinical development and pre-IND studies.